Reticulocyte Maturation Process — Experimental Demonstration of RET Channel Using Anemic Mice —
Sysmex Journal International Vol. 17 Suppl. 1/No. 1 (2007) Reticulocyte Maturation Process — Experimental Demonstration of RET Channel Using Anemic Mice — Mari KONO, Yuri TAKAGI, Tamiaki KONDO, Takuji HANAMURA and Keiji FUJIMOTO Cell analysis center, Scientific Affairs, Sysmex Corporation, 4-4-4 Takatsukadai, Nishi-ku, Kobe 651-2271, Japan Tel.: +81-78-991-1919; Fax: +81-78-991-1942. The RET channel on the XT2000i and XE series of Sysmex automated blood analyzers is a reticulocyte measurement channel. By staining the residual RNA with the florescent dye "RETSEARCH II" it divides reticulocytes into three fractions, high-, medium- and low- fluorescence intensity reticulocytes (HFR, MFR, LFR respectively). Counting the cells is a useful tool for understanding the status of erythropoiesis and is widely used in the diagnosis of for example anemia and bone marrow suppression. It is believed that these fractions represent reticulocytes of different maturation stages, ranging form very early and immature (HFR) to more mature reticulocytes (LFR), but individual features of the different subpopulations have not been characterized to date. Our objectives were to isolate cells of the different stages and to analyze them for differences in morphology and RETSEARCH II staining pattern. Furthermore, we want to compare this to the expression of the well known reticulocyte marker CD71 (Transferrin Receptor) and the new methylene blue staining and classification system. We induced anemia in mice through intraperitoneal phenylhydrazine injection, which leads to severe reticulocytosis in the animals after five days. Anemic murine blood was collected and fractionated into reticulocyte subpopulations by a cell sorting system. The cells were fixed and examined by transmission electron microscopy (TEM) or stained and examined for specific morphology and antigen expression.
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